Information on EC 2.1.1.140 - (S)-coclaurine-N-methyltransferase

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The expected taxonomic range for this enzyme is: Eukaryota, Bacteria

EC NUMBER
COMMENTARY hide
2.1.1.140
-
RECOMMENDED NAME
GeneOntology No.
(S)-coclaurine-N-methyltransferase
REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
S-adenosyl-L-methionine + (S)-coclaurine = S-adenosyl-L-homocysteine + (S)-N-methylcoclaurine
show the reaction diagram
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-
-
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REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
transfer of methyl group
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-
-
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PATHWAY
BRENDA Link
KEGG Link
MetaCyc Link
(S)-reticuline biosynthesis I
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Isoquinoline alkaloid biosynthesis
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Metabolic pathways
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Biosynthesis of secondary metabolites
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SYSTEMATIC NAME
IUBMB Comments
S-adenosyl-L-methionine:(S)-coclaurine-N-methyltransferase
The enzyme is specific for the (S)-isomer of coclaurine. Norcoclaurine can also act as an acceptor.
CAS REGISTRY NUMBER
COMMENTARY hide
167398-06-3
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ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
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-
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Manually annotated by BRENDA team
Berberis koetineana
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-
-
Manually annotated by BRENDA team
Bocconia cordata
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-
-
Manually annotated by BRENDA team
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-
-
Manually annotated by BRENDA team
Corydalis vaginans
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-
-
Manually annotated by BRENDA team
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-
-
Manually annotated by BRENDA team
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-
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Manually annotated by BRENDA team
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-
-
Manually annotated by BRENDA team
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-
-
Manually annotated by BRENDA team
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-
-
Manually annotated by BRENDA team
Thalictrum glaucum
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-
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Manually annotated by BRENDA team
Tinospora fragosa
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-
-
Manually annotated by BRENDA team
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
malfunction
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plants exhibiting reduced CNMT transcript levels show a significant fourfold increase in the relative abundance of papaverine, and a reduction in the accumulation of thebaine and noscapine. Norlaudanine is only detected in CNMT-suppressed plants. The levels of several N-methylated compounds, including N-methylcoclaurine, N,N-dimethylcoclaurine, 4'-O-methyl-N-methylcoclaurine, reticuline, N,N-dimethylreticuline, codamine, laudanine, laudanosine and salutaridine, and four phathelideisoquinoline alkaloids (i.e. narcotoline, noscapine, N,N-dimethylnarcotine and another noscapine-like compound) are also reduced in CNMT-suppressed plants
metabolism
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the enzyme is involved in the biosynthesis pathways of alkaloids in Papaver somniferum, overview. Subsequent 6-O-methylation, N-methylation, 3'-hydroxylation and 4'-O-methylation convert (S)-norcoclaurine to (S)-reticuline, which is a branch-point intermediate in the formation of most BIA structural sub-groups, including morphinan (e.g. morphine), phthalideisoquinoline (e.g. noscapine) and benzophenanthridine (e.g. sanguinarine) alkaloids
SUBSTRATE
PRODUCT                       
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
(R)-coclaurine + S-adenosyl-L-methionine
S-adenosyl-L-homocysteine + (R)-N-methylcoclaurine
show the reaction diagram
-
-
-
?
(R,S)-6-O-methylnorlaudanosoline + S-adenosyl-L-methionine
S-adenosyl-L-homocysteine + (R,S)-N-methyl-6-O-methyl-norlaudanosoline
show the reaction diagram
-
-
-
?
(R,S)-norlaudanosoline + S-adenosyl-L-methionine
S-adenosyl-L-homocysteine + (R,S)-N-methylnorlaudanosoline
show the reaction diagram
-
-
-
?
(R,S)-norreticuline + S-adenosyl-L-methionine
S-adenosyl-L-homocysteine + (R,S)-N-methylnorreticuline
show the reaction diagram
-
-
-
?
(S)-coclaurine + S-adenosyl-L-methionine
S-adenosyl-L-homocysteine + (S)-N-methylcoclaurine
show the reaction diagram
(S)-norcoclaurine + S-adenosyl-L-methionine
S-adenosyl-L-homocysteine + (R)-N-methylnorcoclaurine
show the reaction diagram
-
strictly stereospecific, only (S)-norcoclaurine is N-methylated
-
?
1-methyl-6,7-dimethoxy-1,2,3,4-tetrahydroisoquinoline + S-adenosyl-L-methionine
N-methyl-1-methyl-6,7-dimethoxy-1,2,3,4-tetrahydroisoquinoline + S-adenosyl-L-methionine
show the reaction diagram
-
-
-
?
6,7-dimethoxy-1,2,3,4-tetrahydroisoquinoline + S-adenosyl-L-methionine
N-methyl-6,7-dimethoxy-1,2,3,4-tetrahydroisoquinoline + S-adenosyl-L-methionine
show the reaction diagram
-
-
-
?
S-adenosyl-L-methionine + (R)-coclaurine
S-adenosyl-L-homocysteine + (R)-N-methylcoclaurine
show the reaction diagram
-
122% of the activity with (S)-coclaurine
-
-
?
S-adenosyl-L-methionine + (R)-coclaurine
S-adenosyl-L-homocysteine + (S)-N-methylcoclaurine
show the reaction diagram
-
240% of the activity with (R,S)-norreticuline
-
-
?
S-adenosyl-L-methionine + (R,S)-6-O-methylnorlaudanosoline
S-adenosyl-L-homocysteine + (R,S)-6-O-methyllaudanosoline
show the reaction diagram
-
-
-
?
S-adenosyl-L-methionine + (R,S)-6-O-methylnorlaudanosoline
S-adenosyl-L-homocysteine + (R,S)-N-methyl-6-O-methylnorlaudanosoline
show the reaction diagram
S-adenosyl-L-methionine + (R,S)-norlaudanosoline
S-adenosyl-L-homocysteine + (R,S)-N-methylnorlaudanosoline
show the reaction diagram
S-adenosyl-L-methionine + (R,S)-norreticuline
S-adenosyl-L-homocysteine + (R,S)-N-methylnorreticuline
show the reaction diagram
-
55% of the activity with (S)-coclaurine
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-
?
S-Adenosyl-L-methionine + (S)-coclaurine
?
show the reaction diagram
S-adenosyl-L-methionine + (S)-coclaurine
S-adenosyl-L-homocysteine + (S)-N-methylcoclaurine
show the reaction diagram
S-Adenosyl-L-methionine + (S)-coclaurine
S-Adenosyl-L-homocysteine + N-methylcoclaurine
show the reaction diagram
S-adenosyl-L-methionine + 1-methyl-6,7-dihydroxy-1,2,3,4-tetrahydroisoquinoline
S-adenosyl-L-homocysteine + 6,7-dihydroxy-1,N-dimethyl-1,2,3,4-tetrahydroisoquinoline
show the reaction diagram
S-adenosyl-L-methionine + 6,7-dimethoxy-1,2,3,4-tetrahydroisoquinoline
S-adenosyl-L-homocysteine + 6,7-dimethoxy-N-methyl-1,2,3,4-tetrahydroisoquinoline
show the reaction diagram
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39% of the activity with (S)-coclaurine
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-
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S-adenosyl-L-methionine + 6,7-dimethoxy-N-methyl-1,2,3,4-tetrahydroisoquinoline
S-adenosyl-L-homocysteine + 6,7-dimethoxy-1,2,3,4-tetrahydroisoquinoline
show the reaction diagram
-
180% of the activity with (R,S)-norreticuline
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-
?
additional information
?
-
NATURAL SUBSTRATES
NATURAL PRODUCTS
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
REVERSIBILITY
r=reversible
ir=irreversible
?=not specified
(S)-coclaurine + S-adenosyl-L-methionine
S-adenosyl-L-homocysteine + (S)-N-methylcoclaurine
show the reaction diagram
S-Adenosyl-L-methionine + (S)-coclaurine
?
show the reaction diagram
S-adenosyl-L-methionine + (S)-coclaurine
S-adenosyl-L-homocysteine + (S)-N-methylcoclaurine
show the reaction diagram
COFACTOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
S-adenosyl-L-methionine
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-
METALS and IONS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
Ca2+
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25% increase in activity at 5 mM Ca2+; 5 mM, 25% increase in activity
additional information
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no metal ion requirement
INHIBITORS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
S-adenosyl-L-methionine
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50% inhibition at 0.062 mM, substrate (S)-coclaurine
additional information
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no inhibition by PCMB, iodoacetamide; not inhibited by EDTA, iodoacetamide, p-chloromercuribenzoate
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KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.16 - 0.38
(R,S)-norreticuline
0.04
(S)-Coclaurine
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-
0.05
(S)-norcoclaurine
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probably allosteric enzyme, Hill coeffizient 2.6
0.04 - 0.65
S-adenosyl-L-methionine
Ki VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.062
S-adenosyl-L-methionine
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substrate (S)-coclaurine
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
0.000096
0.0001
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enzyme extract of cell culture, (S)-coclaurine, (R)coclaurine is not methylated
0.0003
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enzyme extract of cell culture, (S)-coclaurine, (R)coclaurine is not methylated
0.00065
Thalictrum glaucum
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enzyme extract of cell culture, (S)-coclaurine, (R)coclaurine is not methylated
0.00066
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enzyme extract of cell culture, (S)-coclaurine, (R)coclaurine is not methylated
0.0007
Corydalis vaginans
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enzyme extract of cell culture, (S)-coclaurine, (R)coclaurine is not methylated
0.00075
Bocconia cordata
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enzyme extract of cell culture, (S)-coclaurine, (R)coclaurine is not methylated
0.00077
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enzyme extract of cell culture, (S)-coclaurine, (R)coclaurine is not methylated
0.0008
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enzyme extract of cell culture, (S)-coclaurine
0.0013
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enzyme extract of cell culture, (S)-coclaurine, (R)coclaurine is not methylated
0.003
Tinospora fragosa
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enzyme extract of cell culture, (S)-coclaurine, (R)coclaurine is not methylated
0.0036
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enzyme extract of cell culture, (S)-coclaurine
additional information
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
8.6
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narrow optimum
pH RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
6 - 9
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about 50% of maximal activity at pH 6.0 and 9.0
6.5 - 8
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about 50% of maximal activity at pH 6.5 and at pH 8.0
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
34 - 37
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23fold purified enzyme preparation, pH 9.0
pI VALUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
4.2
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chromatofocusing
SOURCE TISSUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
SOURCE
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sieve elements of the phloem adjacent or proximal to laticifers. Sieve elements in the root and hypocotyl of opium poppy seedlings
Manually annotated by BRENDA team
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
44000
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recombinant protein, gel filtration and SDS-PAGE
83000
-
gel filtration HPLC
87000
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gel filtration
160000
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gel filtration
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
homotetramer
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alpha, 4 * 45000, SDS-PAGE
tetramer
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4 * 45000, SDS-PAGE
Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
40% ammonium sulfate, Phenyl Sepharose, TSK Fraktogel, Mono Q, Affinity gel, disc electrophoresis, 115fold purification
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; 30-50% ammonium sulfate, Phenyl Sepharose, Q-Sepharose, Mono Q, Mono P, 343fold purification
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recombinant enzyme
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recombinant protein, gel filtration
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Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
expressed in Escherichia coli
expressed in Escherichia coli from the Salmonella enterica prpBCDE promoter, comparison to expression products derived from the strongest IPTG-inducible promoter PT7, DH10B strain used for DNA-manipulations, BLRDE3 strain host for pPro and T7 expression systems
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expressed in Escherichia coli, recombinant CNMT protein
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expression in Escherichia coli
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isolation of molecular clones
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real-time quantitative PCR analysis of target gene transcript levels, comparison of target gene transcript levels and papaverine content in individual opium poppy plants infiltrated with Agrobacterium tumefaciens harboring control and gene-specific pTRV2 constructs
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ENGINEERING
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
additional information
-
virus-induced gene silencing in opium poppy to functionally characterize six genes potentially involved in papaverine biosynthesis, e.g. the gene encoding coclaurine N-methyltransferase, altered alkaloid content phenotypes, overview. Norlaudanine is only detected in CNMT-suppressed plants
APPLICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
biotechnology
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CNMT should be quite useful for biotransformation of the intermediates of alkaloid biosynthesis to N-methyltransferred products